期刊
JOURNAL OF DAIRY SCIENCE
卷 105, 期 8, 页码 6499-6512出版社
ELSEVIER SCIENCE INC
DOI: 10.3168/jds.2021-21409
关键词
thermophilus; CRISPR interference; multiplex gene repression; exopolysaccharide biosynthesis; uridine diphosphate glucose sugar metabolism; Streptococcus thermophilus
资金
- National Natural Science Foundation of China (Beijing, China) [31871776, 31972101]
- National Science Fund for Distinguished Young Scholars (Beijing, China) [32025029]
- Natural Science Foundation of Shanghai (Shanghai, China) [22ZR1444000]
- Shanghai Agriculture Applied Technology Development Program (Shanghai, China) [2019-02-08-00-07- F01152]
- Shanghai Engineering Research Center of Food Microbiology (Shanghai, China) [19DZ2281100]
This study developed a CRISPRi system for gene transcriptional modulation in S. thermophilus. By modulating gene expression in the EPS synthesis module, the production of EPS was increased.
Streptococcus thermophilus is used extensively in the dairy industry and has shown great promise as a chas-sis cell for the biosynthesis of high-value metabolites. However, metabolic engineering in S. thermophilus lacks effective genetic modification tools to modu-late gene expression to relieve metabolic burden and maximize the production of desired compounds. Here, we developed a clustered regularly interspaced short palindromic repeats interference (CRISPRi) system for efficient gene transcriptional modulation in S. ther-mophilus. Our CRISPRi system typically achieved 66 to 98% knockdown of single or multiple gene expres-sion. We used CRISPRi for the biosynthesis of a new exopolysaccharide (EPS) as a paradigm model. Repres-sion of galK at module of uridine diphosphate glucose sugar metabolism and overexpression of epsA and epsE at EPS synthesis module resulted in an approximately 2-fold increase in EPS titer (277 mg/L) when com-pared with a control strain. This study demonstrated the effectiveness of CRISPRi as a powerful metabolic engineering tool and synthetic biology strategy for S. thermophilus.
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